Genotyping Mosquito Populations using SSR Loci

Genomic DNA was individually extracted from 120 mosquitoes (15 males and 15 females for each population) and genotyped at 11 polymorphic simple sequence repeats (SSR) loci: Aealbmic1, 2, 3, 5, 6, 9, 11, 14, 15, 16 and 17 as described in^{51 (link)}. PCR amplifications and fragment identifications were performed as described in^{52 (link)}. Each PCR product was then diluted 1:10 in ddH2O water and 2 μL of this dilution was added to 10 μL of a mixture of deionized formamide and GeneScan-500 ROX size standard (Applied Biosystems, CA, USA). Genotyping was processed in an ABI3730XL sequence analyser (Applied Biosystems) and data analysed using GeneScan and Genemapper software. The first step of the genetic analysis was to bin the genotyped raw data using TANDEM V. 1.09^{53 (link)}; this program helps to overcome problems related with genotyping errors. When microsatellite amplification was not successful or scoring was uncertain, re-extraction of DNA was performed.